摘要
【目的】水稻(Oryza sativa L.)是中国最重要的粮食作物,也是盐胁迫敏感作物。研究水稻盐胁迫响应基因表达,发掘水稻耐盐基因对水稻抵御盐胁迫的分子机制及解析其调控网络,为培育耐盐水稻品种奠定基础。【方法】以水稻品种日本晴种子为试验材料,在1/2MS培养基培养条件下对其进行盐处理(150 mmol·L-1 NaCl),对盐处理和非盐处理21 d的根系进行小分子量RNA组学测序。通过生物信息学分析,以log2 Fold Change(log2FC)>1或<-1为筛选条件,寻找在盐胁迫和非盐胁迫条件下差异表达的miRNA和tRF(tRNA-derived RNA fragments),分析其靶基因,并通过实时荧光定量PCR对测序结果及靶基因进行验证。【结果】以至少一组数据RPM(Reads Per Million reads)>500和log2FC>1或<-1为筛选条件,共得到31个差异表达miRNA,其中8个为盐胁迫诱导miRNA,23个为盐胁迫抑制miRNA,这些差异表达miRNA属于12个miRNA家族,其中8个miRNA家族在拟南芥、玉米和小麦等物种中也被报道为盐胁迫信号响应miRNA,包括盐胁迫抑制的osa-miR397、osa-miR396、osa-miR156、osa-miR167、osa-miR1432和盐胁迫诱导的osa-miR159、osa-miR168、osa-miR164。其余4个miRNA家族osa-miR1882、osa-miR1876、osa-miR1423和osa-miR5077尚未见与盐胁迫相关的报道。通过靶基因预测,得到这31个差异表达miRNA的靶基因共162个。此外,盐处理后,水稻根系产生的34—38nt tRF数量显著多于非盐处理材料,说明tRF的产生并非随机,而是通过某种特定机制响应高盐信号,诱发tRNA特异性加工而产生。以RPM>50和log2FC>1或<-1为筛选条件,检测到盐胁迫诱导的5'端tRF 3个,3'端tRF 3个,这些tRF由6个tRNA加工产生。推测这些差异tRF是潜在的水稻盐胁迫响应tRF。【结论】共检测到12种水稻根系中盐胁迫响应miRNA,其靶基因多为转录因子编码基因,推测其通过对其靶基因转录因子的转录后调控参与了盐胁迫响应的表达调控。其中8个水
【Objective】Rice(Oryza sativa L.)is the most important food crop in China.But as a salt sensitive plant,to study the expression pattern of salt stress responding genes in rice has theoretical and practical significance.The dissection of the molecular mechanism and regulatory network in response to salt stress will contribute to salt-tolerant rice breeding.【Method】The rice variety Nipponbare seeds were used as experimental materials.Samples were treated with 150 mmol·L-1 NaCl for 21 days in 1/2MS medium.Small RNAs were extracted from treated and non-treaded root samples for sequencing.Differentially expressed miRNAs and tRFs(tRNA-derived RNA fragments)were identified using the cut-off of log2 fold change(log2FC)>1 or<-1,and their predicted target genes were analyzed.The sequencing results and target genes were further verified by real-time quantitative PCRs.【Result】RPM>500 for at least one set of data and log2 FC>1 or<-1 were used as the cut-off threshold,and 31 differently expressed miRNAs were obtained.Among them,the expression of 8 miRNA were up-regulated and 23 miRNA were down-regulated under salt treatment.The differently expressed miRNAs belonged to 12 miRNA families.8 of these miRNA families have been reported as salt responding ones in Arabidopsis,maize and wheat,including osa-miR397,osa-miR396,osa-miR156,osa-miR167,osa-miR1432,which were down-regulated miRNAs,and osa-miR159,osa-miR168,osa-miR164,which were up-regulated ones under salt treatment,The other 4 miRNA families,namely osa-miR1882,osa-miR1876,osa-miR1423 and osa-miR5077,have not yet been reported in relation to salt stress.162 target genes of these 31 differentially expressed miRNAs were predicted.Under salt stress,the expression of 34-38nt tRFs were significantly enhanced in rice root,indicating these tRFs were not produced randomly,but induced by salt stress.RPM>50 and log2 FC>1 or<-1 were used as the threshold,3 tRFs were detected as 5'terminal tRFs and 3 tRFs were detected as 3'terminal tRFs.These tRFs are produced from 6 tRNAs.It
作者
孟淑君
张雪海
王琪月
张稳
黄力
丁冬
汤继华
MENG ShuJun;ZHANG XueHai;WANG QiYue;ZHANG Wen;HUANG Li;DING Dong;TANG JiHua(College of Agronomy,Henan Agricultural University/National Key Laboratory of Wheat and Maize Crop Science,Zhengzhou 450002)
出处
《中国农业科学》
CAS
CSCD
北大核心
2020年第4期669-682,共14页
Scientia Agricultura Sinica
基金
河南省高等学校重点科研项目（13A210466）
国家科技重大专项（转基因专项课题）（2018ZX0800908B）
河南省科技攻关项目（182102110349）
河南农业大学科技创新基金（KJCX2019A01）
